Overall Center Abstract This application describes the Pediatric Ohio-New York Cancer (Peds-ONC) Immunotherapy Center, created in response to the RFA as a second nidus for the Pediatric Immunotherapy Discovery and Development Consortium (PI-DDN). To complement the funded U54 (Maris and Mackall, MPI) of the PI-DDN that largely seeks to harness adaptive immunity to further develop CAR-T cells against newly identified antigens, we propose to harness innate immunity to target pediatric cancers, to circumvent resistance to conventional therapy, and to further enable adaptive/hybrid immune approaches. Our aims are to (1) Identify and overcome barriers to utilizing NK and CAR-NK cells as cancer therapeutics, (2) break tolerance to ?self? cancer-associated proteins and (3) enhance immunotherapies by targeting suppressive myeloid cells. We will accomplish our aims through four projects, based at two major sites based in Ohio (Nationwide Children?s Hospital) and New York (New York Medical College) with subsites in Columbus (The Ohio State University) and Minneapolis (Univeristy of Minnesota). In addition to an Administrative Shared Resource (Core A, directed by Dr. Timothy Cripe and Associate Director Dr. Mitchell Cairo), the projects are scientifically supported by a comprehensive Genomics & Immune Monitoring Shared Resource (Core B, directed by Dr. Elaine Mardis with several subspecialy assistant directors). Core B provides integrated datasets via state- of-the-art technologies including mass cytometry, single cell transcriptomics, and an array of other genomics approaches that enable detailed characterizations and tracking of cancer cell immunogenicity, the tumor immune microenvironment, and immunologic responses. We have also assembled strong external and internal scientific advisory boards of renowned leaders whose expertise spans the projects and shared resources. The projects are highly integrated and cross-informative. We propose that therapeutic regimens that combine modalities will produce synergy that drives anti-tumor immune responses in preclinical pediatric cancer models, overcoming the limitations of low mutational burdens. Our goal is to generate a sufficient body of knowledge with compelling data to inform the rational design of future clinical trials and thereby improve the lives of children with cancer. 1